Asthma and allergy are marked by a profound dysregulation of Th2 responses and increased expression of the Th2 cytokines IL5, IL13 and IL4. Genetic variants in the Th2 cytokine locus are strongly associated with allergic inflammation, but their impact on the phenotype is far from understood because the genetic heterogeneity and complexity existing within outbred populations prevent association studies from revealing which genetic variants are involved in phenotype determination. Our goal is to characterize the mechanism(s) underlying the impact of natural variants on the expression and function of genes critical for the development of, and the susceptibility to, human allergic inflammation. Our model gene is IL13, a key effector in asthma and allergy. We showed that: (1) IL13 variants are strong determinants of asthma/allergy;(2) the IL13 locus includes two blocks of highly linked single nucleotide polymorphisms (SNP);and (3) IL13+2044G>A in the 3'block, results in the expression of an IL13 R130Q variant more active than wild type (WT) IL13, whereas IL13-1112C>T in the 5'block increases IL13 transcription in Th2 cells and IL13 secretion ex vivo. Despite these advances, we are convinced ultimately regulatory SNPs need to be studied in vivo within a physiologic genomic context. Here we wish to explore the hypothesis that the impact of genetic variation on human IL13 regulation can be effectively modeled and dissected in BAC transgenic (TG) mice carrying WT or asthma/allergy-associated human IL13 haplotypes. More specifically, we propose: Specific Aim 1: To generate and validate [murine IL13-deficient] BAC TG mouse lines carrying the WT human IL13 locus or the IL13 haplotypes most commonly associated with asthma traits in Caucasians. This work will capitalize on our analysis of variation in IL13 and our skills in BAC recombineering, and will lead to the generation of TG lines exhibiting faithful tissue-specific and copy number-dependent expression of human IL13, but lacking murine IL13. Specific Aim 2: To identify functional IL13 polymorphisms, and the underlying mechanisms, by comparing and contrasting human IL13 regulation and IL13-dependent lung responses in [murine IL13-deficient] BAC TG lines carrying distinct IL13 haplotypes. This work will determine whether variation acts on IL13 transcriptional and/or post-transcriptional regulation or through the secretion of an overactive IL13 protein variant (IL13 R130Q). [The murine IL13-deficient background will be ideal for the characterization of human IL13-dependent phenotypes]. By providing a controlled genetic background, this in vivo model will define the SNPs (or blocks thereof) involved in IL13 dysregulation and their modifying effects on IL13 expression and/or function, paving the way for successful strategies to neutralize the effects of genetic dysregulation in IL13-mediated disease. PUBLIC HEALTH RELEVANCE. The overall goal of our work is to characterize the mechanisms underlying the impact of natural genetic variation on the expression and function of genes critical for the development of, and the susceptibility to, human allergic inflammation. Our model is IL13, a Th2 cytokine which is overexpressed in patients with asthma and/or allergy. We have studied variation in IL13 quite extensively, and we have identified several polymorphisms that dysregulate the expression and/or the function of this gene in vitro. Clearly, a genetically determined increase in IL13 expression and/or activity is likely to play a major role in the pathogenesis of asthma and allergy. Despite these advances, we are convinced novel, more powerful and physiologic approaches are required to elucidate the role played by genetic variation in IL13 dysregulation and IL13- mediated disease. Ultimately, polymorphisms need to be studied in vivo within the physiologic genomic context. In this proposal we wish to explore the hypothesis that (a) human IL13 polymorphisms associated with asthma-related phenotypes are sufficient to induce appreciable dysregulation of IL13 expression and/or function, and (b) the impact of natural genetic variation on human IL13 regulation can be effectively modeled and dissected in mouse models carrying defined wild type or asthma/allergy-associated human IL13 haplotypes [on a murine IL13- deficient background]. By ensuring haplotype-specific patterns of IL13 regulation are gauged against a controlled genetic background, this in vivo model will allow us to determine which polymorphisms are necessary and sufficient for IL13 dysregulation, leading to the molecular mechanisms responsible for altered IL13 expression and/or function.